Coordination to divalent cations by calcium-binding proteins

Abstract

Fourier-transform infrared spectroscopy (FTIR) is a powerful tool for examining the metal coordination of the side chain COO − groups of Glu and Asp on Ca 2+ -binding proteins in solution. The behavior of COO − symmetric stretch can be investigated by using protein samples in H 2 O solution. However, it is difficult to obtain information about the behavior of the COO − antisymmetric stretch in H 2 O solution, because the COO − antisymmetric stretching band overlaps with the amide II band. Therefore, to obtain reliable infrared spectra in the region of COO − antisymmetric stretch, exchangeable protons in the protein should be completely deuterated by incubating the apoprotein dissolved in D 2 O under mild heating conditions.